Next-Generation Sequencing Analysis of CpG Methylation of a Tumor Suppressor Gene SHP-1 Promoter in Stable Cell Lines and HCV-Positive Patients

Viruses. 2022 Oct 26;14(11):2352. doi: 10.3390/v14112352.


Hepatitis C virus (HCV) is the major causative pathogen associated with hepatocellular carcinoma and liver cirrhosis. The main virion component, the Core (C) protein, is involved in multiple aspects of HCV pathology including oncogenesis and immune evasion. In this study, we established a next-generation bisulfite sequencing (NGS-BS) protocol to analyze the CpG methylation profile at the tumor suppressor gene SHP-1 P2 promoter as a model system. Our data show that HCV C protein expression in the immortalized T cells correlated with a specific CpG methylation profile at the SHP-1 P2. The NGS-BS on HCV-positive (HCV+) patient-derived PBMCs revealed a considerably different CpG methylation profile compared to the HCV C protein immortalized T cells. Notably, the CpG methylation profile was very similar in healthy and HCV+ PBMCs, suggesting that the SHP-1 P2 CpG methylation profile is not altered in the HCV+ individuals. Collectively, the NGS-BS is a highly sensitive method that can be used to quantitatively characterize the CpG methylation status at the level of individual CpG position and also allows the characterization of cis-acting effects on epigenetic regulation.

Keywords: CpG methylation; HCV; SHP-1; next-generation sequencing.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Carcinoma, Hepatocellular* / genetics
  • Carcinoma, Hepatocellular* / metabolism
  • Cell Line
  • CpG Islands
  • DNA Methylation
  • Epigenesis, Genetic
  • Genes, Tumor Suppressor
  • Hepatitis C* / genetics
  • High-Throughput Nucleotide Sequencing
  • Humans
  • Liver Neoplasms* / genetics
  • Liver Neoplasms* / metabolism


  • PTPN6 protein, human
  • hydrogen sulfite

Grants and funding

This research was funded by PLP-CM program, SciLife (VC-2022-0005, A.B.), Uppsala County Council (AS19023/42301, A.B.) and a Ph.D. fellowship from European Community Mobility Program Erasmus Mundus Action 2 Namaste (EMA2, P.D.).